JPH0999067A - Method for adsorbing beta 2 microglobulin in body fluid - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a method for adsorbing β2 microglobulin having safety which selectively adsorbs the β2 microglobulin with high efficiency, less adsorbs the albumin in particular and does not require supplementary fluid. SOLUTION: This method for adsorbing the β2 microglobulin in the body fluid is executed by bringing blood or plasma into contact with a porous material having at least surface area of >=10m<2> /g an having the average pore size of the pores ranging from 20 to 2000Å.

Description

Detailed Description of the Invention

[0001]

TECHNICAL FIELD The present invention relates to an adsorption method for selectively adsorbing and removing a malignant substance associated with a disease from body fluids such as blood, plasma, serum, ascites, and pleural effusion. More specifically, β ₂ microglobulin increases in the body fluids of patients with renal failure and malignant tumors and causes carpal tunnel syndrome, amyloidosis, elastic finger / shoulder / knee joint disease, pruritus dermatitis, bone disorders, etc. The present invention relates to an adsorption method for selectively adsorbing and removing.

[0002]

2. Description of the Related Art Approximately 1 patient undergoing hemodialysis in renal failure
After 0 years, abnormalities such as carpal tunnel syndrome have become apparent. In recent years, it has been clarified that this causative substance is β ₂ microglobulin, which is relatively difficult to remove by dialysis, and various symptoms are caused by its accumulation in the body.

Conventionally, hemofiltration and diafiltration have been used for the purpose of removing such a medium molecular weight substance, but the removal rate is low, and there is a problem that a large amount of replacement fluid is required for effective removal. In order to increase the removal rate, it is sufficient to make the pores of the membrane larger, but when the pores become a little larger, albumin, which is a useful protein, is leaked, and controlling the pore size cannot achieve effective selective removal of medium-molecular weight substances. is the current situation.

[0004]

The object of the present invention is generally applicable in view of the problems based on the above-mentioned therapeutic polymer membrane technology, and the medium molecular weight substance, especially β ₂ microglobulin is high. Adsorbs selectively with high efficiency, non-specific adsorption,
Particularly, the present invention intends to provide an adsorption method suitable for purification or regeneration of body fluids, which is less adsorbed by albumin, requires no replenisher, is safe, and can be easily sterilized.

[0005]

Means for Solving the Problems The inventors of the present invention have made extensive studies in accordance with the above-mentioned object, and as a result, have found that a specific porous material selectively adsorbs β ₂ microglobulin with surprisingly high efficiency. I found it. As a result of further detailed examination of this, it was found that β ₂ microglobulin is selectively adsorbed with high efficiency only when the relationship between the average pore diameter of the pores and the surface area satisfies certain conditions. It was completed.

That is, according to the present invention, the surface area is at least 10 m ² / g or more, and the average pore diameter is 20Å.
Is a method for adsorbing β ₂ microglobulin in a body fluid, which is characterized in that blood or plasma is brought into contact with a porous material in the range from 1 to 2000Å.

In the present invention, β ₂ microglobulin is
It is β ₂ microglobulin which is usually measured by enzyme immunoassay in clinical tests, but more specifically, it has the following physical properties. Sedimentation constant 1.6S Partial specific volume 0.72-0.73 ml / g Molecular weight 11000-12000 Nitrogen content 16-17%

The β ₂ microglobulin which is the object of the present invention includes β ₂ microglobulin itself and a complex with other proteins, and also includes a partially altered amino acid sequence of β ₂ microglobulin.

The surface area of the porous material is a value measured by the BET method using nitrogen gas (for example, Catalyst Engineering Course-4, Catalyst Basic Measurement Method, Catalysis Society of Japan, Jijijinkan, pages 50 to 58). refers to an, it must be 10 m ² / g or more, and an average pore diameter of the pores should be in the range of 20Å to 2000 Å. The average pore diameter of the fine pores can be obtained by calculation from the mercury intrusion curve obtained by the mercury intrusion method (pages 69 to 73). The larger the surface area, the better the adsorption efficiency of β ₂ microglobulin, but the range must have a mechanical strength that can maintain the pore size and structurally maintain a porous structure.

If it is less than 10 m ² / g, the capacity as an adsorbent is insufficient. If the average pore size is smaller than 20Å, the adsorption of β ₂ microglobulin becomes extremely poor. That is, as the pore size becomes smaller, the surface area of the porous substance tends to increase, but since β ₂ microglobulin cannot enter inside the pores of the porous material, the substantial surface area for β ₂ microglobulin becomes extremely large. It becomes smaller and the adsorption of β ₂ microglobulin becomes worse. On the other hand, when the pore size is larger than 2000 Å, β ₂ microglobulin can enter the inside of the pores, but the surface area of the porous material becomes small, and the β ₂ microglobulin adsorption efficiency also deteriorates.

On the other hand, the average pore size is 20Å to 200
Within the range of 0Å, β ₂ microglobulin can freely enter inside the pores, and the substantial surface area capable of adsorbing β ₂ microglobulin is sufficiently large, which enables specific and efficient adsorption of β ₂ microglobulin. Become.

Here, the surface area and the average pore diameter are in the range of 10 m ² / g or more and the average pore diameter is 20Å to 2000Å, and more preferably the surface area is 55 m.
² / g or more and an average pore size of 45Å to 1000
It is in the range of Å. More preferably, the surface area is 100 m
² / g or more, and an average pore size of 100Å to 500
It is in the range of Å.

The shape of the porous material used in the present invention is not particularly limited, and any of granular, spherical, flat membrane, hollow fiber, cylindrical, rod-shaped, etc. can be used. Considering the ease of handling and the ease of handling during use, it is preferable that the particles have a particle size of 25 μm to 2500 μm. More preferable particle size is 50 μm to 200
0 μm, particularly preferably 250 μm to 1500
μm.

As the material of the porous material, inorganic materials such as silica, alumina, magnesia, silica-magnesia, silica-alumina and glass, and organic polymer materials are used. The present invention was made by discovering that β ₂ microglobulin is extremely adsorbed on the surface of a foreign material, but from the viewpoint of safety and adsorption affinity of eluate as a body fluid purification material, organic polymer Materials are preferably used.

The organic polymer material preferably has a contact angle of 20 ° or more, more preferably 30 ° or more, and particularly preferably 4 because of its adsorption affinity with β ₂ microglobulin.
Materials of 0 degrees or more are used.

The contact angle referred to in the present invention is the contact angle of air bubbles on the surface of a solid in water, and WC Hamilton, J. Col.
loid Interface Sci., 40, 219-222 (197)
2) [Double Sea Hamilton, Journal of
Colloid Interface Science, 40, 219
-222 (1972)], JDAndrade, J.Polym.Sci.Po.
lym.Symp., 66, 313-336 (1979) [JAD Andrade, Journal of Polymer]
Science Polymer Symposium, 66, 313-
336 (1979)],
Says the measured contact angle. According to the well-known method for measuring the contact angle of a droplet on a solid surface in air, the water-absorbent material has a contact angle value that changes with the passage of time and is difficult to use as a physical property value of the material. . As the sample, a sheet and a film-shaped molded product were prepared, and the contact angle was measured at a temperature of 25.
The temperature was set to 0 ° C., and the measurement was performed 10 times or more.

Preferred organic polymer materials include polyolefin compounds such as polyethylene, polypropylene and polytetrafluoroethylene, polymers of vinyl compounds such as polystyrene, polymethacrylate ester and polyacrylate ester, and polyamides such as nylon 6 and 66. Examples thereof include polyester compounds and polyester compounds such as polyethylene terephthalate. Of these, homopolymers such as methacrylate esters, acrylate esters, ethylene and styrene derivatives, or copolymers with comonomers and crosslinking agents are preferably used. In particular, crosslinked polymer particles containing methyl methacrylate or styrene as a main component are preferably used. As the cross-linking agent, any known cross-linking agent may be used, and examples thereof include divinylbenzene, ethylene glycol di (meth) acrylate, and polyethylene glycol di (meth) acrylate.

In addition to these organic polymer materials, first-class, second-class,
Anion exchangers having a tertiary, tertiary, or quaternary nitrogen atom introduced therein are preferred because they have increased affinity with β ₂ microglobulin.

When the present invention is used for purification of whole blood, it is preferable to coat it for the purpose of preventing the generation of fine particles. Examples of the coating agent include (meth) acrylic acid ester-based polymers, acrylamide-based polymers, polyvinylpyrrolidone-based polymers, polyvinylalcohol-based polymers, cellulose nitrate, and gelatin.

For the purpose of preventing the generation of fine particles, further improving blood compatibility, and further improving the affinity for β ₂ microglobulin, a polymer having a nitrogen-containing basic functional group is most preferable as a coating agent. Used.

The "nitrogen-containing basic functional group" in the present invention is a functional group which has a positive charge on the nitrogen atom in an acidic aqueous solution and can be a cation. Examples of such functional groups include primary amino groups, secondary amino groups, tertiary amino groups, quaternary ammonium groups, and nitrogen-containing aromatic ring groups such as pyridyl groups and imidazolinyl groups.

Accordingly, examples of the polymer having a nitrogen-containing basic functional group used in the present invention include vinylamine;
2-vinylpyridine, 4-vinylpyridine, 2-methyl-5-vinylpyridine, 4-vinylimidazole, N-
Vinyl derivatives of nitrogen-containing aromatic ring compounds such as vinyl-2-ethylimidazole and N-vinyl-2-methylimidazole; dimethylaminoethyl (meth) acrylate, diethylaminoethyl (meth) acrylate, dimethylaminopropyl (meth) acrylate, 3 Acrylic acid and methacrylic acid derivatives such as dimethylamino-2-hydroxypropyl (meth) acrylate; acrylic acid amides such as N-dimethylaminoethyl (meth) acrylic acid amide and N-diethylaminoethyl (meth) acrylic acid amide; Polymerization containing a methacrylic acid amide derivative; a styrene derivative such as p-dimethylaminomethylstyrene and p-diethylaminoethylstyrene; and a derivative obtained by converting the above vinyl compound into a quaternary ammonium salt by using an alkyl halide or the like. And the like.

Of these, particularly preferred are polymers containing dimethylaminoethyl (meth) acrylate, diethylaminoethyl (meth) acrylate, p-dimethylaminomethylstyrene, p-diethylaminoethylstyrene and the like.

Further, the polymer having a nitrogen-containing basic functional group referred to in the present invention is preferably a copolymer of a vinyl compound and a monomer having a nitrogen-containing basic functional group, and the nitrogen content in the functional group. Is preferably 0.05 to 3.5% by weight. The nitrogen content referred to here is the weight% of the nitrogen atoms in the above functional groups in the total polymer.

Examples of the vinyl compound used herein include alkyl (meth) acrylates such as 2-hydroxyethyl methacrylate, methyl (meth) acrylate, ethyl (meth) acrylate, n-butyl (meth) acrylate, and (meth) acrylamide. , N-methyl (meth) acrylamide and other amides, N-vinylpyrrolidone, vinyl acetate, styrene and the like.

Further, as a copolymer of a vinyl compound and a monomer having a nitrogen-containing basic functional group, a block copolymer, a graft copolymer, a random copolymer or the like is used. In the case of a united or block copolymer, it is preferable to have a microdomain structure having an average length of 100Å to 100 µ.

The adsorbent used in the present invention may be used alone, or may be mixed or laminated with other adsorbents. Examples of other adsorbents include activated carbon and the like used in adsorptive artificial kidneys. As a result, a broader clinical effect due to the synergistic effect of the adsorbent can be expected. The adsorbent volume is 50 to 600 ml when used for extracorporeal circulation.
The degree is appropriate.

When the adsorbent is used in the extracorporeal circulation, there are roughly the following two methods. One is to separate the blood taken out from the body into a plasma component and a blood cell component by using a centrifuge or a membrane-type plasma separator, and then pass the plasma component through the device for purification and It is a method of returning it to the body together with the components, and the other is a method of passing blood taken out of the body directly to the device for purification.

Regarding the passage speed of blood or plasma, since the adsorbent has a very high adsorption efficiency, the particle size of the adsorbent can be made coarse and the packing degree can be lowered, so that the shape of the adsorbent layer can be reduced. It is possible to give a high passing speed regardless of how. Therefore, a large amount of body fluid can be treated. As a method for passing the bodily fluid, it may be continuously passed or intermittently used depending on the clinical need or the equipment condition of the facility.

[0030] As described above, the present invention is highly convenient and safe because it adsorbs and removes β ₂ microglobulin in a body fluid with high efficiency and specificity. INDUSTRIAL APPLICABILITY The present invention can be used for general usage of purifying and regenerating body fluids such as autologous blood and plasma, and β ₂ accumulated in body fluids in renal failure, malignant tumor, etc.
It is effective, safe and reliable for the adsorption and removal of microglobulin.

[0031]

EXAMPLES Hereinafter, embodiments of the present invention will be described in more detail with reference to examples. Example 1 Methyl methacrylate-divinylbenzene copolymer particles (80:20 wt%) were used as an adsorbent. (Surface area: 380 m ² / g, average pore size: 160Å, contact angle: 6
5 ± 3 degrees, particle size: 350-500 μm)

After mixing the plasma of the renal failure patient and the adsorbent at a volume ratio of 6: 1, the mixture was shaken at 37 ° C. for 1 hour, and β _{2 in} the plasma before and after was mixed.
Microglobulin and albumin were established. The RIA method was used for β ₂ microglobulin, and the BCG method was used for albumin. The amount of β ₂ microglobulin, which was 44.8 mg / l, was lowered to 1.6 mg / l. On the other hand, albumin only changed from 6.6 g / dl to 6.2 g / dl, and the decrease was slight.

Example 2 Methyl methacrylate-divinylbenzene copolymer particles (80:20 wt%) were used as an adsorbent. (Surface area: 260 m ² / g, average pore size: 400Å, contact angle: 6
5 ± 3 degrees, particle diameter: 350-420 μm) Others were the same as in Example 1.

The previous value of β ₂ microglobulin is 44.8 m
What was g / l decreased to 0.8 mg / l after adsorption. In contrast, 6.6 g / dl of albumin is 6.4.
It changed to g / dl, and the decrease was slight.

Example 3 Commercially available XAD-8 was used as an adsorbent. (ROHM
(Manufactured by Anders Co., Ltd.) (Surface area: 150 m ² / g, average pore size: 180 Å, contact angle: 62 ± 4 degrees, particle size: 350 to 500 μm) Others were the same as in Example 1.

The previous value of β ₂ microglobulin is 44.8 m
What was g / l decreased to 2.0 mg / l. In contrast, albumin 6.6 g / dl is 6.5 g / dl
, And the decrease was slight.

Example 4 Styrene-divinylbenzene copolymer particles (80:20 wt%) were used as an adsorbent. (Surface area: 500 m ² / g, average pore size: 400 Å, contact angle: 76 ± 5, particle size: 350 to 500 μm) Others were the same as in Example 1.

Β ₂ microglobulin has a pre-value of 44.8 m
What was g / l decreased to 0.6 mg / l. On the other hand, albumin 6.6 g / dl is 6.3 g / dl
It just dropped to.

Claims (1)

[Claims] 1. β in a body fluid, which is characterized in that blood or plasma is brought into contact with a porous material having a surface area of at least 10 m ² / g or more and an average pore diameter of 20 Å to 2000 Å. ₂ Microglobulin adsorption method.